Experimental evaluation of Hingusauvarchaladi Ghrita and Saptavartita Hingusauvarchaladi Ghrita with special reference to their anticonvulsant activity

Abstract

Incidence of epilepsy is 0.3 to 0.5% in different populations throughout the world, and the prevalence of epilepsy has been estimated at 5 to 10 persons per 1000. Scanning of the Ayurvedic classics reveals that 90% of the formulations mentioned to have action on sajnavaha srotas are ghrita-based formulations. Ghrita because of its yogavahi guna, incorporates the qualities of the drugs added to it without losing its own qualities. In the present study Hingusauvarchaladi ghrita and saptavartita Hingusauvarchaladi ghrita have been selected, to prove their anticonvulsant activity experimentally on albino mice, by the chemoshock method. The observations recorded have been analyzed by one-way ANOVA followed by Scheffe's test, statistically. Saptavartita Hingusauvarchaladi ghrita has shown better anticonvulsant activity in comparison to Hingusauvarchaladi ghrita.

Introduction

Convulsion is a sign of panic, which is commonly observed in epileptic and non-epileptic conditions. It often causes transient impairment of consciousness, leaving the individual at risk and is quite often seen in epilepsy. The pathophysiological theories of epilepsy have identified the brain as the site of the problem, and in the mid nineteenth century, the pathological changes in the brain associated with epilepsy were recognized.

As per Ayurvedic classical texts Akshepa is a separate disease classified under Vatananatmaja vyadhi[1] as well as it has been found as a symptom in different diseases, such as, Apasmara, Apatanaka, Apatantraka[2] and so on, but it is mainly observed in Apasmara. In Madava Nidana the samprapthi of Akshepa is clearly mentioned. The Ayurvedic treatise has described the causes, symptoms, and treatment of the disease in detail. In spite of many synthetic drugs introduced in the treatment of convulsions, they develop adverse drug reactions like CNS depression, ataxia, sedation, incoordination, thrombocytopenia, pancreatitis, hepatic dysfunction, tetratogenicity, and so forth.[3] There is still a need for an ideal anticonvulsant medicine with properties like broad spectrum activity, rapid onset of action, very few side effects, good bio-availability, cost effective, and so on.

Some of the drugs and therapies mentioned in Ayurvedic literature have been found to be very effective and offer great hope for a satisfactory treatment of this disease. Today in the era of scientific advances, it is necessary to validate these drugs and therapies experimentally and clinically as well.

Here an attempt has been made to find out the effect of Hingusauvarchaladi ghrita and Sapthavartita Hingusauvarchaladi ghrita experimentally, to prove their anticonvulsant activity.

Aims and Objectives

• To revalidate the anticonvulsant activity of the trial drugs, experimentally.

• To compare the anticonvulsant activity of the trial drugs.

• To evaluate the anticonvulsant activity of the trial drugs in comparison to the standard drug sodium valproate.

Materials and Methods

Drugs

The raw drugs necessary for the present study were collected from the local market at Udupi and were identified as genuine samples by the Department of Dravyaguna, A.L.N. Rao Memorial Ayurvedic Medical College Koppa. The entire practical was done in the pharmacy of the Bhaishajya Kalpana Department, A.L.N. Rao Memorial Ayurvedic Medical College, Koppa.

Trial drug 1-Hingusauvarchaladi ghrita:

Trial drug 2 - Saptavartita hingusauvarchaladi ghrita

The process was repeated seven times with the same ingredients of trial drug 1. Concept of avartana was adopted.

Experimental study

Selection of animals

Twenty-four albino mice of either sex, weighing between 20 and 25 g, bred in an animal house (Reg. No. 191/CPCSEA) of A.L.N. Rao Memorial Ayurvedic Medical College and P.G. Center were selected for the study, after getting clearance from the Institutional Ethical Committee. Six mice were taken in each group. The animals were housed in groups under 12-hour light / dark cycles, with standard diet and water ad libitum, to avoid the ‘kindling’ effect that occurred as a result of repeated convulsions induced by experimental methods. The animals were not subjected to any experiments for about seven days. The dose was calculated as per the table of Paget and Barne's [Table 1].

Table 1.

Grouping of animals

Methodology-The chemoshock method by pentylenetetrazol

Pentylenetetrazol[4] is a CNS stimulant. It produces jerky type of clonic convulsions in rats and mice. The convulsive effect of this drug is considered to be analogous with the petitmal type of convulsions in man. In recent times Pentylenetetrazol has been reported to act through GABA – a benzodiazepine receptor mechanism in the brain. It is widely used as a tool in experimental pharmacology to study the convulsive and anticonvulsive actions of the drugs.

Drugs

Pentylenetetrazol (dose 60 mg/kg, i.p)

A stock solution containing 6 mg/ml of the drug was prepared and 1 ml / 100 g of body weight of the mouse was injected. A dose of PTZ 0.25 ml / 25 g body weight of mouse was injected intraperitonially.

The following parameters were observed and recorded as time in seconds:

Seizure duration

• Clonus

• Tonic hind limb flexion

• Tonic hind limb extension

• Postictal depression

• Recovery / Death

Control group

Animals in this group were administered distilled water 0.1 ml orally, and after an interval of 60 minutes, pentylenetetrazol (60 mg/kg body weight) was injected intraperitonially and the different phases of seizures were noted.

Standard group

All the animals in this group were administered sodium valproate 0.32 ml mixed orally and after 60 minutes, pentylenetetrazol (60 mg/kg body weight) was injected intraperitonially to observe the effects of the drug.

Trial group – T1

These animals were administered hingusauvarchaladi ghrita for three days at a dose of 0.16 ml orally and on the third day, after administration of the drug, an interval of 60 minutes was maintained, and then they were subjected to chemical shock by injecting pentylenetetrazol in a dose of 60 mg/kg body weight, intraperitonially. The results obtained were recorded.

Trial Group-T2

These animals were administered saptavartita hingusauvarchaladi ghrita for three days at a dose of 0.16 ml orally, and on the third day after administration of the drug, an interval of 60 minutes was maintained, and then they were subjected to chemical shock by injecting pentylenetetrazol in a dose of 60 mg/kg body weight, intraperitonially. The results obtained were recorded.

Results

The mean time duration of clonus in the control group, trial T-1 and T-2 was 13.17, 8.5 and 7.16 seconds respectively and abolition of clonus was noted in standard group. In the control group all animals exhibited tonic hind limb flexion, the mean time duration was 9.5 seconds. In standard group except 4, all animals have shown abolition of tonic hind limb flexion and the mean time duration was 2.17 seconds. In trial T- 2 there was abolition of the symptom in one mice .In the control group, all animals exhibited tonic hind limb extension and the mean time duration was 9.17 seconds. In the standard group, all animals except in 3 abolition of tonic hind limb extension was found and the mean time duration was 1.16 seconds. In T-1 and T-2 abolition of tonic hind limb extension was observed in 1 animal and 2 animals respectively [Table 2].

Table 2.

P.T.Z. induced convulsions mean values and standard error mean (parameters are recorded as time in seconds)

Discussion

In Ayurvedic texts Akshepa (convulsion) is a separate disease, classified under Vatananatmaja Vyadhi as well as it has been found as a lakshana in different diseases like Apasmara, Apatanaka, Apatantraka, and so on, but it is mainly observed in Apasmara. The trial drug Hingusauvarchaladi ghrita is used as medicine for Apasmara. With this background an attempt has been made to prepare Hingusauvarchaladi ghrita and saptavartita Hingusauvarchaladi ghrita and to find out their anticonvulsant activity. In this method, the criterion noted for evaluation of the anticonvulsant activities of the control, standard, and trial compounds is abolition of the clonus, although criteria such as the tonic hind limb flexion and tonic hind limb extension have also been noted as useful guidelines to assay anticonvulsant activity.

The standard drug and the trial drug T1 and T2 have produced significant (P < 0.05) reduction in the seizure duration when compared to the control [Table 2].However, the standard drug abolished the clonus phase, and the trial drugs T1 and T2 significantly reduced its duration when compared with the control group. The standard drug caused a significant reduction in the extensor--flexor ratio (E/F) when compared to the control group. The trial drug 2 also had a significantly reduced E/F ratio. The trial drugs also showed significantly reduced postictal depression when compared to the control group.

From the above results it was concluded that both the trial drugs possessed anticonvulsant activity. On comparing trial drug 1 and trial drug 2, it was concluded that the trial drug 2 possessed better anticonvulsant activity than trial drug 1 [Table 2].

Recovery / death of animals

Death noted in groups C, T1, and T2, four, three, and two, respectively. The remaining animals recovered in all the respective groups. In the standard group all the animals recovered. From this result it could be concluded that trial drug 2 possessed better anticonvulsive activity as compared to trial drug 1.

Probable mode of action

The probable mode of action of the trial drugs can be explained in four angles. Hingu, sauvarchala, trikatu, and gomutra are ushna veerya and vatakapha shamaka drugs.

Akshepaka is a vata-predominant disease, where the sanjavaha srotas are blocked and the patient goes into an unconscious condition. Here hingu is the best sapjna sthapana drug[5], thereby it acts as an Akshepa hara dravya. Trikatu helps to increase the bioavailability of the medicine and moreover removes the srotorodha in the cellular level. Hingu and sauvarchala[6] lavana are the best Anulomana, Deepana, and pachana dravyas, which help to subdue the aggravated vata. The kshareeya guna of gomutra and sauvarchala lavana are best to remove the srotorodha. In this compound formulation hingu acts as the activator, gomutra and sauvarchala lavana act as the potentiators, trikatu acts as the bioavailability enhancer, and ghee being the base of the formulation increases the shelf life of the product.

As the active ingredients are mixed with ghee they easily get digested and absorbed. The lipophilic action of the ghee facilitates transportation to the target site.

In general, the blood cerebrospinal fluid and the blood brain barriers are highly permeable to water, carbon dioxide, oxygen, and most lipid soluble substances, and slightly permeable to electrolytes such as sodium chloride and potassium. The more lipophilic the drug is, the more likely it is to cross the blood-brain barrier [7][8].The active principles of hingu, sauvarchala lavana, trikatu, and gomutra are lipid soluble, and hence, they cross the blood-brain barrier, to bring about the sapjna sthapana of the afflicted person.

Conclusion

The trial drugs 1 and 2, that is, Hingusauvarchaladi ghrita and saptavartita hingusauvarchaladi ghrita, have a definite demonstrable anticonvulsant action as ascribed by the experimental study conducted on albino mice. The trial drug 1 — Hingusauvarchaladi ghrita and trial drug 2 — saptavartita Hingusauvarchaladi ghrita have a moderately significant effect against convulsions, when compared with the standard drug. The trial drug 2 — saptavartita Hingusauvarchaladi ghrita was more effective than the trial drug 1 — Hingusauvarchaladi ghrita.